1. Field of The Invention
The invention relates to devices and methods for biomolecule purification.
2. Description of The Prior Arts
Gel filtration chromatography and conventional preparative gel electrophoresis are two of the most popular means employed for biomolecule purifications.
In gel filtration chromatography, a buffer flows through a matrix in a column. A sample of biomolecule mixture applied over the matrix is carried across the matrix by the flow of the buffer. There are numerous pores existing on the beads of the matrix. The separation of biomolecules relies on movement of the biomolecules into and out of the pores. The biomolecules at sizes larger than that of the pores can not enter the pores and move rapidly across the matrix. The biomolecules at sizes smaller than that of the pores enter and leave the pores repeatedly and therefore retain longer in the matrix. A separation between a group of large biomolecules and a group of small ones is then achievable by collecting them into separated fractions.
In conventional preparative gel electrophoresis, buffer does not flow through a gel matrix. But all biomolecules have to travel across the pores of the gel matrix. Movement of biomolecules is driven by an interaction between a net charge of the biomolecules and an electric potential applied on the gel matrix. The migration rate of a given biomolecule in the gel matrix is determined by its size, its shape, its net charge, the pore size of the gel matrix, and the potential difference of the electric potential. Thus different biomolecules in a mixture can be distinctly separated from each other at high resolution by sequentially collecting eluted fractions from a preparative gel apparatus. The migration rate of biomolecules in conventional preparative gel electrophoresis is much slower than that in gel filtration chromatography so that an extended process of fraction collection is unavoidable during electrophoresis. Thus an external fraction collector is essentially required.
Gel filtration chromatography devices and methods gain their popularity in routine biomolecule purifications by their speed and convenience. Pharmacia Biotech Inc. describes, for example, a series of pre-packed disposable gel filtration columns in its catalog of 1995 as a handy tool for such applications. These devices and methods, however, fail to achieve further purification whenever a higher resolution is required.
Conventional preparative gel electrophoresis devices and methods are utilized for high demanding biomolecule purifications when high resolution is a critical issue. Dorman U.S. Pat. No. 3,539,493 teaches, for example, a device and method of conventional preparative gel electrophoresis. It requires an installation of a special ion-permeable glass disc 108, an external fraction collector connecting to a elution tubing 70, two external mechanical pumps connecting to buffer inlet 94 and port 88 respectively, and a complicated procedure of assembly and disassembly of the device for each application. Hediger U.S. Pat. No. 4,479,861 teaches a device and method of conventional preparative gel electrophoresis. It requires an installation of a dialysis membrane, an external fraction collector, two external mechanical pumps, and a complicated procedure of assembly and disassembly for each application as well. Caccavo U.S. Pat. No. 3,980,546 teaches a device and method of conventional preparative gel electrophoresis. An external fraction collector and two external mechanical pumps, while a dialysis membrane is omitted, are still required for its function. Huynh U.S. Pat. No. 5,151,165 teaches a device and method of conventional preparative gel electrophoresis. To omit the requirement of using external mechanical pump, a delicate installation of an elution chamber is introduced. Besides, an external fraction collector and a dialysis membrane are still unavoidable. Similar examples can also be found at Chen U.S. Pat. No. 4,877,510 and Lim et al U.S. Pat. No. 5,284,559. The advantage of high resolution of these devices and methods, however, exhibits no attraction in applications of routine biomolecule purifications due to their complicated setup procedure, their high labor tensity, their tedious cleanup process, and their requirements of using an external fraction collector, using external pumps, and using a large bench space. The feasibility of these devices and methods is even ended in applications in which the material to be purified is radioactive or is at an extremely low amount level.
None of existing devices and methods own both high resolution and convenience together simultaneously. But, there are numerous routine biomolecule purifications in which both high resolution and handy to use are highly desired.